1. Field of the Invention
The invention relates to the fields of microbiology and molecular biology. In particular, the invention relates to the field of mycology and the production of secondary metabolites from fungi.
2. Summary of the Related Art
Secondary metabolites are a major source of commercially useful products such as food additives, vitamins, and medicines for the treatment of a wide variety of infections and diseases. By way of example, in 1997 the statin drugs lovastatin, simvastatin, and pravastatin, fungal secondary metabolites used in the treatment of hypercholesteremia, together had US sales of US$7.53 billion (Sutherland et al., Current Opinion In Drug Discovery & Development 4:229-236 (2001)). The cost and availability of these plant, bacterial and fungal metabolites are frequently determined by limitations imposed on production and purification of these compounds from culture. This problem is frequently exacerbated by the fact that these products are generally produced during the stationary phase of bacterial and fungal growth.
A wide variety of methods have been utilized to increase the amount of secondary metabolite produced in culture. Studies have demonstrated the importance of carefully designing the medium in which a fungus is grown to maximize the amount of a secondary metabolite produced (see, e.g., Hajjaj H, et al., Appl. Environ. Microbiol. 67:2596-602 (2001); Lesova, K., et al., J. Basic Microbiol. 40:369-75 (2000)). In addition, the method of culture or fermentation also impacts directly on the amount of secondary metabolite produced. For example, see Robinson, T., et al. (Appl. Microbiol. Biotechnol. 55:284-289 (2001)), which demonstrates the advantages of solid state (substrate) fermentation.
In addition to the manipulation of culture and media conditions, genetic approaches have been taken to increase secondary metabolite production. For example, the production of penicillin is limited by the activity of two enzymes, encoded by the ipnA and acvA genes, both of which are regulated by the pacC protein, a zinc-finger transcription factor. Naturally occurring mutant alleles of the pacC locus are known to possess more transcription-activating activity than the cognate, wild-type allele (see, e.g., Tilburn et al. EMBO J. 14(4):779-790 (1995)). Thus, one genetic approach to increasing secondary metabolite production is to identify and isolate naturally occurring mutant alleles, the expression of which leads to increased secondary metabolite production.
Although many regulators of secondary metabolite production in many organisms are known, not all of the organisms that produce secondary metabolites are amenable to genetic or molecular genetic manipulation. Thus, these systems are not generally useful as a source for the isolation of naturally occurring mutant alleles and are even less useful for the deliberate manipulation of secondary metabolite regulator protein structure with the aim of creating improved regulators of secondary metabolite production.
It would be advantageous to have improved regulators of the biosynthetic enzymes responsible for secondary metabolite production. For example, recent studies suggest increasing usage of statin drugs, e.g., see Waters D. D., Am. J. Cardiol. 88:10F-5F (2001)). Thus, demand for statin drugs is likely to increase substantially. In order to meet the demand for these and other secondary metabolites, new and improved methods for the production of secondary metabolites must be identified.